Project Summary: The NIH has committed substantial resources to both traditional and - through its Protein Structure Initiative - high-throughput approaches to protein structure determination, and similar commitments have been made in Europe and Asia. A survey of the results for soluble proteins shows that the main bottlenecks are obtaining high-quality crystals and obtaining high-quality X-ray diffraction data for accurate structures. The long-term goals of this project are to understand the underlying science involved in macromolecular crystal growth, in cryopreservation of protein crystals, and in radiation damage during X-ray data collection, and to use this understanding to develop practical protocols and technologies. These studies utilize a wide array of physical methods, from synchrotron-based X-ray diffraction and imaging to microfabrication. Our specific goals are: (1) To understand the fundamental processes in cryopreservation, and to develop more effective methods to capture and preserve protein and crystal structure based on ultra- fast cooling, the kinetic phase diagrams of aqueous mixtures, and the thermal expansion behavior of solvent and crystal;(2) To. develop methods for cryopreservation and long-term storage of protein solutions that preserve crystallization behavior;(3) To quantify and understand radiation damage in protein crystals, and to develop protocols for reducing this damage in small crystals;and (4) To explore protein drop-surface interactions and drop dynamics relevant to protein crystallization. Relevance: Protein crystallography is a central component of modern structural genomics and drug discovery efforts, which promise to revolutionize medicine in the 21st century. The proposed research will facilitate more rapid and accurate determination of protein structures, and thereby assist in understanding the mechanisms underlying disease and in developing new treatments.